The sustainable utilization of black soldier fly (BSF) for recycling organic waste into nutrient-rich biomass, such as high-quality protein additive, is gaining momentum, and its microbiota is thought to play important roles in these processes. Several studies have characterized the BSF gut microbiota in different substrates and locations; nonetheless, in-depth knowledge on community stability, consistency of member associations, pathogenic associations, and microbe–microbe and host–microbe interactions remains largely elusive. In this study, we characterized the bacterial and fungal communities of BSF larval gut across four untreated substrates (brewers’ spent grain, kitchen food waste, poultry manure, and rabbit manure) using 16S and ITS2 amplicon sequencing. Results demonstrated that substrate impacted larval weight gain from 30 to 100% gain differences among diets and induced an important microbial shift in the gut of BSF larvae: fungal communities were highly substrate dependent with Pichia being the only prevalent genus across 96% of the samples; bacterial communities also varied across diets; nonetheless, we observed six conserved bacterial members in 99.9% of our samples, namely, Dysgonomonas, Morganella, Enterococcus, Pseudomonas, Actinomyces, and Providencia. Among these, Enterococcus was highly correlated with other genera including Morganella and Providencia. Additionally, we showed that diets such as rabbit manure induced a dysbiosis with higher loads of the pathogenic bacteria Campylobacter. Together, this study provides the first comprehensive analysis of bacterial and fungal communities of BSF gut across untreated substrates and highlights conserved members, potential pathogens, and their interactions. This information will contribute to the establishment of safety measures for future processing of BSF larval meals and the creation of legislation to regulate their use in animal feeds.

Gut microbiota plays important roles in many physiological processes of the host including digestion, protection, detoxification, and development of immune responses. The honey bee (Apis mellifera) has emerged as model for gut-microbiota host interaction studies due to its gut microbiota being highly conserved and having a simple composition. A key gap in this model is understanding how the microbiome differs regionally, including sampling from the tropics and in particular from Africa. The African region is important from the perspective of the native diversity of the bees, and differences in landscape and bee management. Here, we characterized the honey bee gut microbiota in sub-Saharan Africa using 16S rRNA amplicon sequencing. We confirm the presence of the core gut microbiota members and highlight different compositions of these communities across regions. We found that bees from the coastal regions harbor a higher relative abundance and diversity on core members. Additionally, we showed that Gilliamella, Snodgrassella, and Frischella dominate in all locations, and that altitude and humidity affect Gilliamella abundance. In contrast, we found that Lactobacillus was less common compared temperate regions of the world. This study is a first comprehensive characterization of the gut microbiota of honey bees from sub-Saharan Africa and underscores the need to study microbiome diversity in other indigenous bee species and regions.

Stingless bees (Apidae: Meliponini) are the most diverse group of corbiculate bees and are important managed and wild pollinators distributed in the tropical and subtropical regions of the globe. However, little is known about their associated beneficial microbes that play major roles in host nutrition, detoxification, growth, activation of immune responses, and protection against pathogens in their sister groups, honeybees and bumble bees. Here, we provide an initial characterization of the gut bacterial microbiota of eight stingless bee species from sub-Saharan Africa using 16S rRNA amplicon sequencing. Our findings revealed that Firmicutes, Actinobacteria, and Proteobacteria were the dominant and conserved phyla across the eight stingless bee species. Additionally, we found significant geographical and host intra-species-specific bacterial diversity. Notably, African strains showed significant phylogenetic clustering when compared with strains from other continents, and each stingless bee species has its own microbial composition with its own dominant bacterial genus. Our results suggest host selective mechanisms maintain distinct gut communities among sympatric species and thus constitute an important resource for future studies on bee health management and host-microbe co-evolution and adaptation.

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Background: Nasopharyngeal samples contain higher quantities of bacterial and host nucleic acids relative to viruses; presenting challenges during virus metagenomics sequencing, which underpins agnostic sequencing protocols. We aimed to develop a viral enrichment protocol for unbiased whole-genome sequencing of respiratory syncytial virus (RSV) from nasopharyngeal samples using the Oxford Nanopore Technology (ONT) MinION platform. Methods: We assessed two protocols using RSV positive samples. Protocol 1 involved physical pre-treatment of samples by centrifugal processing before RNA extraction, while Protocol 2 entailed direct RNA extraction without prior enrichment. Concentrates from Protocol 1 and RNA extracts from Protocol 2 were each divided into two fractions; one was DNase treated while the other was not. RNA was then extracted from both concentrate fractions per sample and RNA from both protocols converted to cDNA, which was then amplified using the tagged Endoh primers through Sequence-Independent Single-Primer Amplification (SISPA) approach, a library prepared, and sequencing done. Statistical significance during analysis was tested using the Wilcoxon signed-rank test. Results: DNase-treated fractions from both protocols recorded significantly reduced host and bacterial contamination unlike the untreated fractions (in each protocol p<0.01). Additionally, DNase treatment after RNA extraction (Protocol 2) enhanced host and bacterial read reduction compared to when done before (Protocol 1). However, neither protocol yielded whole RSV genomes. Sequenced reads mapped to parts of the nucleoprotein (N gene) and polymerase complex (L gene) from Protocol 1 and 2, respectively. Conclusions: DNase treatment was most effective in reducing host and bacterial contamination, but its effectiveness improved if done after RNA extraction than before. We attribute the incomplete genome segments to amplification biases resulting from the use of short length random sequence (6 bases) in tagged Endoh primers. Increasing the length of the random nucleotides from six hexamers to nine or 12 in future studies may reduce the coverage biases.